title: Inhibition of lung tumorigenesis by a small molecule CA170 targeting the immune checkpoint protein VISTA

status: Public on Jun 04 2021

city: Chicago

first: Weiguo

last: Cui

state: IL

address: 303 E Chicago Ave

department: Pathology

institute: Northwestern University

summary: Expressed on cells of the myeloid and lymphoid lineages, V-domain Ig Suppressor of T cell Activation (VISTA) is an emerging target for cancer immunotherapy. Blocking VISTA activates both innate and adaptive immunity to eradicate tumors in mice. Using a tripeptide small molecule antagonist of VISTA CA170, we found that it exhibited potent anticancer efficacy on carcinogen-induced mouse lung tumorigenesis. Remarkably, lung tumor development was almost completely suppressed when CA170 was combined with an MHCII-directed Kras peptide vaccine. Flow cytometry and single-cell RNA sequencing (scRNA-seq) revealed that CA170 increased CD8+ T cell infiltration and enhanced their effector functions by decreasing the tumor infiltration of myeloid-derived suppressor cells (MDSCs) and Regulatory T (Treg) cells, while the Kras vaccine primarily induced expansion of CD4+ effector T cells. VISTA antagonism by CA170 revealed strong efficacy against lung tumorigenesis with broad immunoregulatory functions that influence effector, memory and regulatory T cells, and drives an adaptive T cell tumor-specific immune response that enhances the efficacy of the Kras vaccine.

overall_design: 5-6 week old female A/J mice were divided into five groups: (a) control mice that received PBS; (b) adjuvant control mice that received a STING agonist plusAddaVax; (c) KVax mice treated with KVax plus adjuvant; (d) CA170 treatment (10mg/kg in PBS) by oral gavage five times per week; and (e) combined treatment with KVax/adjuvant plus CA170. The experimental treatment scheme is shown in. Mice were administered one dose of the carcinogen vinyl carbamate30 (VC; Toronto Research Chemicals, Inc.) by i.p. injection (16 mg/kg in sterile saline). CA170 treatment was started one week after VC induction of lung adenocarcinoma. KVax was started two weeks after VC induction, followed by four more vaccinations at 2-week intervals, and additional boost vaccines every four weeks for the duration of the experiments. Primary lung tumors were harvested and pooled from different treatment groups at the end of the study, then minced and digested at 37oC for 20 min with mouse tumor dissociation buffer (MiltenyiBiotec, CA) to generate single cell suspensions per the manufacturer’s instructions. Tumor-infiltrating leukocytes were directly stained with 7-AAD, CD45 and CD3 surface markers, and CD3+ or CD45+ TILs were flow-sorted.

first: Yao

last: Chen

first: Jing

last: Pan

first: Ming

last: You


title: Genome-wide mapping of ChARF3-binding sites in hazel

status: Public on Jun 05 2021

city: Siping

first: Yunqing

last: Cheng

address: Haifeng street

country: China

institute: Jilin Normal University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The goal of this study is to identify direct target of ChARF3 during ovule developmental stages.

overall_design:
ChIP experiment was performed using the specific polyclonal antibody against ChARF3. Three pairs of the input control and antibody-treated ChIP-Seq libraries were constructed using ovules from three developmental stages.
Polyclonal antibodies against a 380 aa long peptide sequence (1-380 aa), representing a unique region in the ChARF3 protein sequence, were raised (Wuhan ABclonal Biotechnology Co., Ltd.).

first: Heng

last: Wei

first: Jianfeng

last: Liu


title: Topological isolation as a mechanism for precise control of developmental regulators in mammalian genomes [RNA-seq]

status: Public on Jun 06 2021

city: Beijing

first: Hua-Jun

last: Wu

state: Beijing

address: 8 science park road, zhongguancun life science park

department: School of Basic Medical Sciences

institute: Peking University Health Science Center

type: Expression profiling by high throughput sequencing

summary: Recent investigation of insulated chromosome structures suggests their crucial role in controlling proper gene expression. However, it remains unclear whether developmental regulators can be precisely controlled by such insulated structures and whether developmental failure is caused by disrupting the structure boundaries. Here we identified topologically insulated regions (TIRs) from deeply sequenced human embryonic stem cell (hESC) Hi-C data and analyzed the gene distribution across TIRs. We find a notable enrichment of developmental regulators isolated from other genes by TIRs and show that the boundary CTCF sites are highly conserved across species. Perturbation of such boundaries in hESCs leads to failure to properly differentiate. Enhancer activity is restricted by such boundaries, further pointing towards their regulatory importance. Finally, germline mutations affecting such boundaries are subject to purifying selection and are underrepresented in the human population. Our findings provide new insights into mammalian genome organization and suggest a possible role of gene insulation through chromosomal folding structures for precise control of developmental regulators.

overall_design: RNA-Seq of SOX17 upstream CTCF wildtype and knockout in embryonic differentiation

first: Alexandro

last: Landshammer

first: Alexander

last: Meissner

first: Franziska

last: Michor


title: Topological isolation as a mechanism for precise control of developmental regulators in mammalian genomes [HiC]

summary: Developmental regulators are spatiotemporal regulated in the process of human development. Recent investigation of insulated chromosome structures suggests their crucial role in control of proper gene expression. However, it’s unclear whether developmental regulators can be precisely controlled by such insulated structures and whether developmental failure can be caused by disrupting the structure boundaries. Here we identified topologically insulated regions (TIRs) from deeply sequenced human embryonic stem cell (hESC) Hi-C data and analyzed the gene distribution across TIRs. We found a notable enrichment of developmental regulators isolated from other genes by TIRs and show that the boundary CTCF sites are highly conserved across species. Perturbation of such boundaries in hESCs leads to the differentiation failure. Germline mutations affecting such boundaries are subject to purifying selection, and are underrepresented in the human population. Further analyses demonstrated the enhancer activity is restricted by such boundaries that further points towards regulatory importance. Our findings provide new insights into mammalian genome organization and including a possible role of gene insulation through chromosomal folding structures for precise control of developmental regulators.

overall_design: HiC-Seq of 4 cell lines


title: Gene expression profiles in SOX10-deficient human iPS cell-derived neural crest cells

city: Changsha

first: Jian

last: Song

state: 湖南

address: 87 Xiangya Road

department: Department of Otorhinolaryngology

institute: Xiangya Hospital Central South University

summary: Waardenburg syndrome (WS) is a autosomal dominant inherited disorder characterized by sensorineural hearing loss (SNHL) and pigment abnormalities. SOX10 is one of its main pathogenicity genes. In vitro generation of patient-specific induced pluripotent stem cells (iPSCs) is an effective approach for exploring the mechanisms of human disease. Here, we established an iPSCs from a WS patient with SOX10 mutation and differentiated it into induced neural crest cells (NCCs), the key cell type involves in inner ear development. Compared with control-derived iPSCs, the SOX10 mutant iPSCs showed significantly decreased efficiency of development and differentiation potential at the stage of NCCs. We then performed high-throughput RNA-seq and examined the transcriptional misregulation at each stage. The transcriptome analysis differentiated NCCs reveals widespread gene expression changes and the differentially expressed genes(DEGs) were enriched with gene ontology (GO) of neuron migration, skeletal system development, multicellular organism development, which indicate the important role of SOX10 plays in the differentiation of NCCs. It's worth noting that, a significant enrichment among the nominal DEGs for genes implicated in inner ear development was found, as well as several genes related to the inner ear morphogenesis. Based on protein-protein interaction（PPI）net work, we selected four candidate genes that may be regulated by SOX10 in inner ear development: BMP2, LGR5, GBX2 and GATA3. In conclusion, the SOX10 deficiency in this WS patient has a significant impact on the gene expression patterns during the development of NCCs. The DEGs most significantly enriched the inner ear development and morphogenesis could help explain the underlying basis for the inner ear malformation seen in WS patient.

overall_design: Gene expression profiles were compared between wild type iPS cell-derived neural crest cells and SOX10 mutant iPS cell-derived neural crest cells

first: Jie

last: Wen

first: Yijiang

last: Bai

first: Yong

last: Feng


title: Gut-derived inflammatory cytokines modulate olfactory perception through metabolic reprogramming of ensheathing glia.

status: Public on Jun 02 2021

city: South San Francisco

first: Jason

middle: A

last: Hackney

state: CA

address: 1 DNA Way

department: Department of Bioinformatics and Computational Biology

country: USA

institute: Genentech, Inc.

summary: Infection-induced aversion against enteropathogens is a conserved sickness behavior that can promote host survival1-3. The etiology of this behavior remains poorly understood, but studies in Drosophila have linked olfactory and gustatory perception to avoidance behaviors against toxic microbes4-6. Whether and how enteric infections directly influence sensory perception to induce or modulate such behaviors remains unknown. Here we show that enteropathogen infection in Drosophila can modulate olfactory perception through metabolic reprogramming of ensheathing glia of the antennal lobe. This reprogramming depends on JAK/STAT signaling activation in ensheathing glia as a consequence of infection-induced Unpaired cytokine expression in the intestine. JAK/STAT signaling induces the expression of glial monocarboxylate transporters (MCTs) and the apolipoprotein glial lazarillo (Glaz), resulting in changes in glial lipid metabolism. This modulates olfaction sensitivity, promoting avoidance of bacteria-laced food and increasing animal survival. While transient in young animals, gut-induced metabolic reprogramming of ensheathing glia becomes constitutive in old animals due to age-related intestinal inflammation, contributing to an age-related decline in olfaction sensitivity. Our findings identify adaptive glial metabolic reprogramming by gut-derived cytokines as a mechanism causing lasting changes in neuronal activity in the aging animal.

overall_design: We have labelled pan glia of Drosophila with tdTomato in the presence of repo::Gal4 driver, and have further purified STAT::GFP+ glia and STAT::GFP- glia correspondingly from mock or infected young flies.

first: Xiaoyu

middle: T

last: Cai

first: Hongjie

last: Li

first: Martin

middle: B

last: Jensen

first: Elie

last: Maksoud

first: Jovencio

last: Borneo

first: Yuxin

last: Liang

first: Stephen

middle: R

last: Quake

first: Liqun

last: Luo

first: Pejmun

last: Haghighi

first: Heinrich

last: Jasper


title: RNA-seq of HUVECs stimulated with HG and oxLDL

city: Nanjing

first: Zhiguang

last: Shi

state: Jiangsu

address: 101Longmian Avenue, Jiangning District

institute: Nanjing Medical University

summary: We investigated  the role of SNO-GNIA2 in HG+oxLDL-induced endothelial inflammation during the development of diabetes-accelerated atherosclerosis and found that SNO-GNAI2 could promote endothelial inflammation through dysregulating Hippo-YAP .  We hypothesized that SNO-GNAI2 induced Hippo-YAP dysfunction through enhancing coupling and activating  G-protein coupling receptors (GPCRs).

overall_design: We determined  the role of SNO-GNIA2 in HG+oxLDL induced endothelial inflammation during the development of diabetes-accelerated atherosclerosis and tried to investigate the underlying mechanism, mRNA profiles of HUVECs stimulate with HG andoxLDL were generated by deep sequencing, in duplicate, using Illumina HiSeq. The sequence reads that passed quality filters were analyzed at the transcript isoform level. Fragments Per Kilobase per Million bases (FPKM) analysis was used to evaluate the level of each gene.

last: Ji

first: Menglin

last: chao

first: Shanshan


title: Pancreatic Ppy-expressing gamma-cells display mixed phenotypic traits and adaptive plasticity to engage insulin production

city: Geneva

first: Marta

last: Perez Frances

state: Geneva

address: 1, rue Michel-Servet

country: Switzerland

institute: University of Geneva

summary: The cellular identity of pancreatic polypeptide (Ppy)-expressing γ-cells, the rarest pancreatic islet cell-type, remains elusive. Within islets, glucagon and somatostatin, released respectively from α- and δ-cells, modulate the secretion of insulin by β-cells. Dysregulation of insulin production raises blood glucose levels, leading to diabetes onset. Here, we present the genetic signature of human and mouse γ-cells. Using newly developed tools, we identified a set of genes and pathways defining their functional identity. We found that the γ-cell population is heterogeneous, with subsets of cells producing another hormone in addition to Ppy. These bihormonal cells share identity markers typical of the other islet cell-types. In mice, Ppy gene inactivation or conditional γ-cell ablation did not alter glycemia nor body weight. Interestingly, upon β-cell injury induction, γ-cells exhibited gene expression changes and some of them engaged insulin production, like α- and δ-cells. In conclusion, we provide a comprehensive characterization of γ-cells and highlight their plasticity and therapeutic potential.

overall_design:
Transcriptomic profile of pancreatic Ppy-expressing cells using single cell and bulk RNA-seq
Profile the transcriptomic landscape of Ppy-expressing cells and compare it to the other pancreatic endocrine cell types.
RNA-seq of sorted alpha, beta, delta and gamma-cells from multiple transgenic mouse models.

**Please note that the alpha and beta-cell data was downloaded from the accession number is GSE155519. All sequencing data were uploaded to, and aligned on the Galaxy project against Ensembl reference genome GRCm38.p6 (release 100) using STAR version 2.7.2b in 2-pass mapping mode. Aligned data were counted using HTSeq version 0.9.1 in union mode.

last: Pérez-Francés

first: Léon

last: van Gurp

first: Maria

middle: V

last: Abate

first: Valentina

last: Cigliola

first: Kenichiro

last: Furuyama

first: Eva

last: Brú-Tarí

first: Daniel

last: Oropeza

first: Taïna

last: Carreaux

first: Yoshio

last: Fujitani

first: Fabrizio

last: thorel

first: Pedro

middle: L

last: Herrera


title: Topological isolation as a mechanism for precise control of developmental regulators in mammalian genomes

first: Hua-Jun

last: Wu

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Identification of 22 novel susceptibility loci associated with testicular germ cell tumors

status: Public on Jun 01 2021

city: Philadelphia

first: Matthew

middle: E.

last: Johnson

state: PA

address: 3615 Civic Center Boulevard, Abramson Research Center, Suite 1102G

department: Genetics

institute: Children's Hospital of Philadelphia


title: Identification of 22 novel susceptibility loci associated with testicular germ cell tumors [ATAC-seq]

summary: To investigate the role of the testicular germ cell tumor cells in complex traits, we generated ATAC-seq and Promoter Focused Capture C to gain insight into the gene regulatory arcitecture contacting promoters in an testicular cell line NT2-D1 model of testicular cancer

overall_design: We performed ATAC-seq 2102EP, NCCIT, TCAM2, NT2-D1 and Promoter Focused Capture C on NT2-D1 cells.

middle: E

first: Struan

middle: F

last: Grant


title: Heterogeneity of meningeal B cells reveals a lymphopoietic niche at the CNS borders [scBCR-seq]

city: St. Louis

first: Marco

last: Colonna

state: MO

address: 660 S. Euclid Ave.

institute: Washington University in St. Louis

summary: The meninges contain adaptive immune cells that provide immunosurveillance of the CNS. These  cells are thought to derive from the systemic circulation. Through single cell analyses, confocal 35 imaging, bone marrow chimeras and parabiosis experiments, we show that meningeal B cells derive locally from the calvaria, which harbors a bone marrow niche for hematopoiesis. B cells reach the meninges from the calvaria through specialized vascular connections. This calvaria- meningeal path of B cell development may provide the CNS with a constant supply of B cells educated by CNS antigens. Conversely, we show that a subset of antigen-experienced B cells that 40 populate the meninges in aging mice are blood-borne. These results identify a private source for  meningeal B cells which may help maintain immune privilege within the CNS.

overall_design: scBCR-seq was performed on bulk preparations from mouse peripheral blood, tibial bone marrow, and dura mater from young and aged mice. Sequencing was completed in two batches: The first batch contained 3 mice from each age:tissue combination. The second batch contained peripheral blood and dura from 3 young and 4 old mice.

first: Simone

last: Brioschi

first: Wei-Le

last: Wang

first: Vincent

last: Peng


title: Heterogeneity of meningeal B cells reveals a lymphopoietic niche at the CNS borders


title: Heterogeneity of meningeal B cells reveals a lymphopoietic niche at the CNS borders [scRNA-seq]

overall_design: scRNA-seq was performed on bulk preparations from mouse peripheral blood, tibial bone marrow, and dura mater from young and aged mice. Sequencing was completed in two batches: The first batch contained 3 mice from each age:tissue combination. The second batch contained peripheral blood and dura from 3 young and 4 old mice.


title: CUT&Tag ChIP-seq analysis of BATF in CX3CR1+ effector , CX3CR1- Ly108- exhausted, or Ly108+  progenitor CD8 T cells during chronic viral infection.

first: Weiguo

last: Cui

summary: During chronic viral infection, pathogen-specifc CD8+ T cells develop into three main phenotypically and functionally distinct subsets:  TCF1hi progenitor,  PD-1hi exhausted, and recently identied CX3CR1+ cytotoxic effector cells. Although genetic programs governing progenitor and exhausted subset formation have been well-studied, how CX3CR1+ effector CD8+ T cell differentiation is transcriptionally and epigentically regulated remains elusive. In this study, our single cell transcriptomics and epigenetic assays revealed that three subsets of virus-specific cells were governed by distinct gene regulatory networks (GRNs) and epigenetic landscapes. Computational analyses demonstrated a striking similarity between the CX3CR1+ subset and short-live effector cells (SLECs) from acute LCMV infection. Consistently, genetic deletion of  T-bet (Tbx21) significantly diminished the formation and function of the CX3CR1+ subset. Importantly, we identify that the transcription factor (TF) BATF is required to maintain a permissive chromatin structure that allows differentiation transition from TCF1+ progenitor to CX3CR1+ effector cells. Intriguingly,  haplodificiency of BATF in CD8+ T cells abolished CX3CR1+ effector subset formation. Lastly, we found that BATF directly bound to key genetic regions such as Tbx21 and modulated their enhancer accessibility to facilitate progenitor to CX3CR1+ effector cell transition. These mechanistic insights can be harnessed to overcome T cell exhaustion in treating chronic infections and cancer.

overall_design: BATF CUT&TAG ChIPseq, 3 samples (CX3CR1+ , CX3CR1- Ly108- PD1+, or Ly108+  CD8 T cells), 3 biological replicates per sample, from LCMV-Cl13 infected C57BL/6 mice.

first: Yao

last: Chen


title: Identification of 22 novel susceptibility loci associated with testicular germ cell tumors [Capture C]

summary: To investigate the role of the testicular germ cell tumor cells in complex traits, we generated ATAC-seq and Promoter Focused Capture C to gain insight into the gene regulatory arcitecture contacting promoters in an ttesticular cell line NT2-D1 model of testicular cancer


title: FVE promotes RNA-directed DNA methylation by facilitating the association of RNA polymerase V with chromatin [H3K27me3 ChIP-Seq].

first: xinjian

last: He

address: No. 7, zhongguancun life science park road, changping district

institute: National Institute of Biological Sciences

summary: DNA methylation can be established by RNA-directed DNA methylation (RdDM) in plants. The association of RNA polymerase V (Pol V) with chromatin is a critical step for RdDM. While the SRA-domain-containing proteins SUVH2 and SUVH9 and the DDR complex are known to be required for the association of Pol V with chromatin, it is unknown whether the association of Pol V with chromatin requires other unidentified regulators. Here we found that SUVH9 is able to interact with a conserved histone-interaction protein, FVE, and a previously uncharacterized RRM domain-containing protein, which we named RRM1. We demonstrated that FVE facilitates the association of Pol V with chromatin and thus contributes to DNA methylation at a substantial subset of RdDM target loci, while RRM1 is only slightly involved in RdDM. FVE-dependent RdDM target loci are more abundant in gene-rich chromosome arms than FVE-independent RdDM target loci. FVE was previously shown to be a shared subunit of the RPD3-type histone deacetylase complex and the polycomb-type histone H3K27 trimethyltransferase complex, both of which are involved in transcriptional repression. This study reveals a previously uncharacterized role of FVE in RdDM and suggests that FVE may coordinate RdDM, histone deacetylation, and H3K27 trimethylation, thus ensuring transcriptional silencing of TEs in gene-rich chromosome arms to protect genes from harmful effects of potentially transcribed TEs

overall_design: Examination of H3K27me3 levels in wild type grown under long day condition.

first: Xin-Jian


title: A human mutation in STAT3 promotes type 1 diabetes through a defect in CD8+ T cell tolerance [scRNA, scTCR]

city: Stanford

first: Julia

last: Belk

state: California

address: 450 Serra Mall

institute: Stanford University

summary: scRNA-seq datasets of immune cells in K392R and WT mice

first: Jeremy

last: Warshauer


title: Genome-wide mapping of m6A modified chromatin associated RNA

city: Hangzhou

first: Yuanhui

last: Mao

state: Zhejiang

address: 1369 West Wenyi Road

institute: Zhejiang University

summary: We sequenced the cytoplasmic and chromatin associated RNAs with and without heat stress in mouse embryonic fibroblasts cells. A chromatin associated lncRNA Heat was found to be transcribed upon heat stress and regulates heat response

overall_design: m6A-seq and RNA-seq of chrmotain associated mRNAs under normal and heatshock conditions

first: Quanquan

last: Ji

first: Xin

last: Zong

first: Shu-Bing

last: Qian


title: Transcriptional profiling analysis of the effects of Ssn3 inhibition in Candida albicans

city: Hanover

first: Deborah

middle: Ann

last: Hogan

state: NH

department: Microbiology and Immunology

institute: Geisel School of Medicine

summary: Genome-wide transcriptional profiles of Candida albicans without and with inhibition of an analog sensitive Ssn3 using 3-MB-PP1.

overall_design: Using Candida albicans with an analog sensitive version of Ssn3, we performed proteomics and transcriptomics analyses of effects of Ssn3 inhibition in medium that does not normally stimulate hyphal growth. Cultures were grown in triplicate and analyzed immediately after inhibition and at 3h after inhibition.

first: Jeffery

last: Hollomon


title: Single-cell transcriptomics reveals immune response of intestinal cell types to viral infection

city: Heidelberg

first: Sergio

last: Triana

address: Meyerhofstr. 1

department: Structural and Computational biology

country: Germany

institute: EMBL

summary: In this work we have employed a single cell sequencing approach using 10x Genomics scRNAseq to study Astroviru infection of of ileum organoids

overall_design: 10x Genomics single-cell RNA sequencing V2 and V3.1  was perfomed on Ileal organoids infected with Astrovirus Infection and Ileum biopsies


title: Spatial Distribution of LTi-like Cells in Intestinal Mucosa Regulates Type 3 Innate Immunity

first: Marco

summary: Lymphoid tissue inducer (LTi)-like cells are tissue resident innate lymphocytes that rapidly secrete cytokines that promote gut epithelial integrity and protect against extracellular bacterial infections. Here we report that the retention of LTi-like cells in conventional solitary intestinal lymphoid structures (SILT) is essential for controlling LTi-like cell function and is maintained by expression of the chemokine receptor CXCR5. Deletion of Cxcr5 functionally unleashed LTi-like cells in a cell intrinsic manner, leading to uncontrolled IL-17 and IL-22 production. The elevated production of IL-22 in Cxcr5-deficient mice improved gut barrier integrity, and protected mice during infection with the opportunistic pathogen Clostridium difficile. Interestingly, Cxcr5-/- mice developed LTi-like cell aggregates that were displaced from their typical niche at the intestinal crypt, and LTi-like cell hyperresponsiveness was associated with the local formation of these unconventional SILT. Thus, LTi-like cell positioning within mucosa controls their activity via niche-specific signals that temper cytokine production during homeostasis

overall_design: RNA-seq was performed on CCR6+ ILC3 isolated from the small intestial lamina propria of wild type and CXCR5 KO mice

first: Jennifer

middle: K

last: Bando

first: Cristiane

last: Secca de Silva


title: Circulating proinflammatory cytokines alter transcriptional activity within fibrotic tissue of Dupuytren’s disease patients

first: Ryan

last: Tomlinson

address: 1015 Walnut Street

department: Orthopaedic Surgery

institute: Thomas Jefferson University

summary: In this project, we have utilized blood and tissue samples from patients with Dupuytren’s disease (DD), a common fibroproliferative disorder of the hand, to identify increased circulating proinflammatory cytokines and assess alterations in the tissue using RNAseq and FTIR analysis. Our results indicate that specific cytokines in circulation play a direct role in local disease progression and may be useful for prospective identification of patients at risk for disease progression or recurrence as well as the development of novel therapeutic strategies. Furthermore, these results help to mechanistically link the well understood association of DD with chronic low-grade inflammatory states.

overall_design: RNA sequencing of tissue from patients with Dupuytren's disease


title: Glucocorticoid receptor collaborates with pioneer factors and AP-1 to execute genome-wide regulation

city: Ithaca

first: Erin

middle: Margaret

last: Wissink

state: NY

address: Biotech Bldg Room 417, 529 Campus Rd

department: Molecular Biology & Genetics

institute: Cornell University

summary: The glucocorticoid receptor (GR) regulates transcription through binding to specific DNA motifs, particularly at enhancers. While the motif to which it binds is constant across cell types, GR has cell-type specific binding at genomic loci, resulting in expression of different genes. The presence of other bound transcription factors (TFs) is hypothesized to specify where GR binds. Here, we addressed the roles of other TFs in the glucocorticoid response by comparing changes in GR binding and nascent transcription at candidate cis-regulatory elements in two distinct cell types. We found that each cell type binds GR at thousands of non-overlapping loci, but only a small fraction of these sites also show changes in transcription. GR binding is associated with pioneer factor binding, whereas transcription is induced at sites bound by AP-1. These results support a model of transcriptional regulation in which multiple classes of TFs are required. The pioneer factors increase chromatin accessibility, facilitating the binding of GR and additional factors. AP-1 then poises a fraction of sites to be rapidly activated upon glucocorticoid-induced GR binding. GR also induces transcription at a specific set of enhancers. The coordinated activity of multiple TFs then results in cell-type specific changes in gene expression. We anticipate that many models of inducible gene expression also require multiple distinct TFs that act at multiple rate-limited steps of transcriptional regulation.

overall_design: Nascent transcription in cells with different dexamethasone treatments.

middle: M

first: John

last: Lis


title: The Cdc42/Rac1 regulator CdGAP is a novel E-cadherin transcriptional co-repressor with Zeb2 in breast cancer

city: Boston

first: Marie-Anne

last: Goyette

address: 450 Brookline avenue

institute: DFCI

summary: The loss of E-cadherin causes dysfunction of the cell-cell junction machinery, which is an initial step in epithelial-to-mesenchymal transition (EMT), facilitating cancer cell invasion and the formation of metastases. A set of transcriptional repressors of E-cadherin (CDH1) gene expression, including Snail1, Snail2 and Zeb2 mediate E-cadherin down-regulation in breast cancer. However, the molecular mechanisms underlying the control of E-cadherin expression in breast cancer progression remain largely unknown. Here, by using global gene expression approaches, we uncover a novel function for Cdc42 GTPase-activating protein (CdGAP) in the regulation of expression of genes involved in EMT. We found that CdGAP used its proline-rich domain to form a functional complex with Zeb2 to mediate the repression of E-cadherin expression in ErbB2-transformed breast cancer cells. Conversely, knockdown of CdGAP expression led to a decrease of the transcriptional repressors Snail1 and Zeb2, and this correlated with an increase in E-cadherin levels, restoration of cell-cell junctions, and epithelial-like morphological changes. In vivo, loss of CdGAP in ErbB2-transformed breast cancer cells impaired tumor growth and suppressed metastasis to lungs. Finally, CdGAP was highly expressed in basal-type breast cancer cells, and its strong expression correlated with poor prognosis in breast cancer patients. Together, these data support a previously unknown nuclear function for CdGAP where it cooperates in a GAP-independent manner with transcriptional repressors to function as a critical modulator of breast cancer through repression of E-cadherin transcription. Targeting Zeb2-CdGAP interactions may represent novel therapeutic opportunities for breast cancer treatment.

overall_design: Differential gene expression profile of ErbB2-expressing breast cancer cells shCTL and shCdGAP  (Illumina HiSEq 2000)

first: Jean-François

last: Côté

first: Nathalie

last: Lamarche


title: FVE promotes RNA-directed DNA methylation by facilitating the association of RNA polymerase V with chromatin

first: xinjian

last: He

type: Methylation profiling by high throughput sequencing


title: Repression of CHROMOMETHYLASE 3 Prevents Epigenetic Collateral Damage in Arabidopsis

city: Wageningen

first: Michael

middle: D

last: Nodine

state: Gelderland

address: Radix West

department: Plant Sciences

country: Netherlands

institute: Wageningen University & Research

summary: DNA methylation has evolved to silence mutagenic transposable elements (TEs) while typically avoiding the targeting of endogenous genes. Mechanisms that prevent DNA methyltransferases from ectopically methylating genes are expected to be of prime importance during periods of dynamic cell cycle activities including plant embryogenesis. However, virtually nothing is known regarding how DNA methyltransferase activities are precisely regulated during embryogenesis to prevent the induction of potentially deleterious and mitotically stable genic epimutations. Here, we report that microRNA-mediated repression of CHROMOMETHYLASE 3 (CMT3) and the chromatin features that CMT3 prefers help prevent ectopic methylation of thousands of genes during embryogenesis that can persist for weeks afterwards. Moreover, CMT3-induced ectopic methylation of genes undergoing transcriptional activation can reduce their corresponding transcript levels. Therefore, the repression of CMT3 prevents epigenetic collateral damage on endogenous genes. We also provide a model that may help reconcile conflicting viewpoints regarding the functions of gene-body methylation that occurs in nearly all flowering plants.

overall_design: Methyl cytosine sequencing libraries were generated from 2-4 biological replicates of bent cotyledon stage embryos or seedlings from four genotypes. mRNA-seq libraries were generated from 9 biological replicates of bent cotyledon embryos from three genotypes.

first: Ranjith

last: Papareddy


title: Pathogenesis, miR-122 gene-regulation, and protective immune responses after acute equine hepacivirus infection

first: Gerlinde

last: Van de Walle

address: 235 Hungerford Hill Road

summary: Equine hepacivirus (EqHV) is phylogenetically the closest relative of hepatitis C virus (HCV) and shares genome organization, hepatotropism, transient or persistent infection outcome, and the ability to cause hepatitis. Thus, EqHV studies are important to understand equine liver disease, and further as an outbred surrogate animal model for HCV pathogenesis and protective immune responses. Here, we aimed to characterize the course of EqHV infection and associated protective immune responses. Approach & Results: Seven horses were experimentally inoculated with EqHV, monitored for 6 months, and rechallenged with the same, and subsequently a heterologous EqHV. Clearance was the primary outcome (6 of 7) and was associated with subclinical hepatitis characterized by lymphocytic infiltrate and individual hepatocyte necrosis. Seroconversion was delayed and antibody titers waned slowly. Clearance of primary infection conferred non-sterilizing immunity resulting in shortened duration of viremia after rechallenge. Peripheral blood mononuclear cell responses in horses were minimal, although EqHV specific T cells were identified. Additionally, an interferon stimulated gene signature was detected in the liver during EqHV infection, similar to acute HCV in humans. EqHV, as HCV, is stimulated by direct binding of the liver-specific microRNA, miR-122. Interestingly, we found that EqHV infection sequesters enough miR-122 to functionally affect gene regulation in the liver. This RNA-based mechanism thus could have consequences for pathology. Conclusions: EqHV infection in horses typically has an acute resolving course, and the protective immune response lasts for at least a year and broadly attenuates subsequent infections. This could have important implications to achieve the primary goal of an HCV vaccine; to prevent chronicity while accepting acute resolving infection after virus exposure.

overall_design: Single cell RNA sequencing of libraries prepared from equine peripheral blood mononuclear cells isolated from a single donor before NPHV infection (pre-infection) and at peak viremia timepoints.

first: Joy

first: Raphael

last: Wolfisberg

first: Ulrik

last: Fahnøe

first: Roosheel

middle: S

last: Patel

first: Sheetal

last: Trivedi

first: Arvind

last: Kumar

first: Himanshu

last: Sharma

first: Louise

last: Nielsen

first: Sean

middle: P

last: McDonough

first: Jens

last: Bukh

first: Bud

middle: C

last: Tennant

first: Amit

last: Kapoor

first: Brad

last: Rosenberg

first: Charles

last: Rice

first: Thomas

middle: J

last: Divers

first: Troels

last: Scheel


title: Transcriptional analysis of liver tissue from indeterminate pediatric acute liver failure patients

first: Matthew

middle: J.

last: Schipma

address: 750 N. Lake Shore Drive

department: Biochemistry and Molecular Genetics

institute: Northwestern University

summary: Background and Aims: Many patients with indeterminate pediatric acute liver failure (PALF) have evidence of T-cell driven immune injury, however the precise inflammatory pathways have not been well defined. We have thus characterized the hepatic cytokine and transcriptional signatures of PALF patients. Approach and Results: Retrospective review was performed on 22 children presenting with indeterminate (IND-PALF; n=17) or other diagnoses (DX-PALF; n=6) with available archived liver tissue. Specimens were stained for CD8 T-cells and scored as dense, moderate, or minimal. Measurement of immune analytes and RNA-sequencing (RNA-seq) were performed on whole liver tissue. Immune analyte data was analyzed by principal component analysis, and RNA-seq was analyzed by unsupervised hierarchical clustering, differential gene expression, and gene set enrichment analysis. Most IND-PALF patients (94%) had dense/moderate CD8 staining and were characterized by Th1 immune analytes including TNFα, IFNγ, IL-1β, IL-12, CXCL9, and CXCL12. Transcriptional analyses identified 2 transcriptional PALF phenotypes. Most patients in Group 1 (91%) had IND-PALF and dense/moderate CD8 staining. This group was characterized by increased expression of genes and cell-subset specific signatures related to innate inflammation, T-cell activation, and antigen stimulation. Group 1 also expressed significantly higher levels of gene signatures for T-regulatory cells, macrophages, Th1 cells, T effector memory cells, cytotoxic T-cells, and activated dendritic cells (p-adj <0.05). In contrast, patients in Group 2 exhibited increased expression for genes involved in metabolic processes. Conclusions: IND-PALF patients have evidence of a Th1 mediated inflammatory response driven by IFNγ. Transcriptional analyses suggest a complex immune network may regulate an immune-driven PALF phenotype with less evidence of metabolic processes. These findings provide novel insight into mechanisms of hepatic injury in PALF, areas for future research, and potential therapeutic targets.

overall_design: RNA sequencing of frozen liver tissue from 22 children with acute liver failure and 2 normal controls

first: Catherine

middle: A

last: Chapin

first: Sarah

last: Taylor

first: Estella

last: Alonso

first: Padmini

last: Malladi

last: Shipma


title: Shutoff of host transcription triggers a toxin-antitoxin system to cleave phage RNA and abort infection [toxIN_T4infection_RNAseq]

city: Cambridge

first: Chantal

last: Guegler

state: MA

address: 31 Ames Street

department: Biology

institute: MIT

summary: The purpose of this experiment was to characterize ToxN activity following T4 infection.

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: Two replicates each of +toxIN (pBR322-toxIN) and -toxIN (pBR322 empty vector) cells i0,2.5,5,10,20, and 30 min post-T4 infection.

first: Michael

middle: T

last: Laub


title: Shutoff of host transcription triggers a toxin-antitoxin system to cleave phage RNA and abort infection [toxIN_T7infection_RNAseq]

summary: The purpose of this experiment was to characterize ToxN activity following T7 infection.

platform_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: Two replicates each of +toxIN (pBR322-toxIN) and -toxIN (pBR322 empty vector) cells i0,2.5,5,10,20, and 30 min post-T7 infection.


title: A mycorrhiza-associated receptor-like kinase with an ancient origin in the green lineage

city: Cologne

first: Tak

last: Lee

address: Carl-von-Linne weg 10

institute: Max Planck Institute for Plant Breeding Research

summary: Receptor-like kinases (RLKs) are key cell signalling components. The rice ARBUSCULAR RECEPTOR-LIKE KINASE 1 (OsARK1) functions during a stage post-arbuscule development in the arbuscular mycorrhizal (AM) association and belongs to an undefined subfamily of RLKs. Our phylogenetic analysis revealed that ARK1 has an ancient paralogue, ARK2. Single ark2 and ark1ark2 double mutants in rice showed a non-redundant AM symbiotic function for OsARK2. Global transcriptomics identified a set of genes co-regulated by the two RLKs suggesting OsARK1 and OsARK2 to orchestrate symbiosis in a common pathway. ARK lineage proteins harbour a newly identified SPARK domain in their extracellular regions which underwent parallel losses in ARK1 and ARK2 in monocots. This protein domain has ancient origins in the green lineage and defines additional overlooked groups of putative cell surface receptors

overall_design: Four different genotypes (ark1-2,ark2,ark1ark2,wild type) of rice were subjected to mycorrhizal fungi R. irregularis. Ten biological replicates were generated for each genotype. To study the effects of mutants ark1 and ark2, the transcriptomes were analyzed against the wild type controls.

first: Héctor

last: Montero


title: Shutoff of host transcription triggers a toxin-antitoxin system to cleave phage RNA and abort infection [toxN_overexpression_RNAseq]

summary: The purpose of this experiment was to characterize ToxN cleavage specificity and locations of cleavage genome-wide in E. coli following overexpression.

overall_design: One replicate of +ToxN and empty vector samples after 5 minutes of induction (rRNA subtracted) in the presence of toxI leaky expression, and one replicate of +ToxN and empty vector samples after 10 minutes of induction without toxI leaky expression (rRNA subtracted)


title: Shutoff of host transcription triggers a toxin-antitoxin system to cleave phage RNA and abort infection [toxIN_riftreatment_RNAseq]

summary: The purpose of this experiment was to characterize ToxN activity following activation by rifampicin treatment.

overall_design: Two replicates each of +toxIN (pBR322-toxIN) and -toxIN (pBR322 empty vector) cells 0,2.5,5,10,20, and 30 min post-rifampicin treatment.


title: Expression data from adult male mouse hypothalamus

city: Bronx

first: Anita

last: Autry

address: 1410 Pelham Parkway S

department: Neuroscience and Psychiatry

institute: Albert Einstein College of Medicine

summary:
Laser captured regions surrounding the perifornical area (PeFA) in adult male mice that attacked pups;  a control outside region (unk); a c-fos positive region (c-fos); an oxytocin positive region (oxt)
In this dataset, we include the expression data obtained from dissected mouse forelimbs using a variety of gain- and loss-of-function hedgehog pathway mutants, as well as limbs dissected into responsive (posterior 2/3ds) and non-responsive (anterior 1/3d) Hh tissues. These data are used to obtain 753 genes that are differentially expressed in response to Shh signaling.

overall_design: We obtained 3 biological replicates containing pooled tissue from 6 males for each specified laser-captured area (n=18 total).  C-fos, oxytocin, and outside regions were obtained from the same samples such that the samples are paired observations (Sample 1, sample 2, sample 3 are the names of the 3 replicates of pooled samples).

first: Ming

last: Tang


title: Clca1 expression in the aged kidney in mice

city: San Antonio

first: Yidong

last: Chen

state: Texas

address: 8403 Floyd Curl Drive, MSC 7784

department: Population Health Sciences

institute: UT Health Science Center at San Antonio

summary: In this study, a discovery approach was designed to identify novel mRNA transcripts that may be involved in kidney aging. RNA-Seq revealed increase in the expression of Clca1 mRNA in the aged kidney. CLCA1 facilitates Cl- current operated by TMEM16A, a Ca++-dependent Cl- channel. Cl- movement accompanied by cations such as Na+ and K+ or in exchange for other anions occurs among fluid compartments in many segments of the nephron. Cl- shift among these compartments is governed by electrochemical potential and transporters and exchangers.

overall_design: Four RNA samples extracted from the kidney cortex of young mice (mean age 5.2 months) were compared to four aged mice (mean age 30.2 months). Differential expression analysis were carried out to identify genes that showed over- or under-expression levels in two age-grouped samples.

first: Hak

first: Balakuntalam

last: Kasinath

first: Zhao

last: Lai


title: Shutoff of host transcription triggers a toxin-antitoxin system to cleave phage RNA and abort infection

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Nitrate-induced gene expression in Arabidopsis mutant of NIN-LIKE PROTEIN 2

city: Bunkoy-ku

first: Shuichi

last: Yanagisawa

state: Tokyo

address: Yayoi 1-1-1

department: Biotechnology Research Center

country: Japan

institute: The University of Tokyo

summary: Plants modulate gene expression profile in response to nitrate through NIN-LIKE PROTEIN (NLP) transcription factors in order to promote nitrate uptake and utilization. Although there are 9 NLP proteins in Arabidopsis, NLP7 had attracted most of the attention. Here, we focused on NLP2, because the nlp2 and nlp7 knockout mutants displayed different phenotypes. To gain insight into genes whose expression was affected by the nlp2 mutation, we compared gene expression profiles in the wild-type Columiba and the nlp2 mutant that were treated with 10 mM KNO3 for 1 hour. Columbia and nlp2 that were treated with 10 mM KCl for 1 hour were used as controls. We found that nitrate induction of 6 genes was compromised only in the nlp2 mutant.

overall_design: The wild-type Columbia and the nlp2 mutant were grown under a nitrate-free condition and treated with 10 mM KNO3 or KCl (control) for 1 hour

first: Mineko

last: Konishi


title: KDM5 inhibition is a novel therapeutic strategy for the treatment of KMT2D mutant lymphomas [2]

city: London

first: James

last: Heward

state: England

address: Haemato Oncology, Barts Cancer Institute, Charterhouse Square

country: United Kingdom

institute: Barts Cancer Institute

summary: Loss of function mutations within KMT2D are a striking feature of the germinal centre lymphomas, with lesions present in 80% of Follicular Lymphoma (FL) and 30% of Diffuse Large B-cell Lymphoma (DLBCL), resulting in decreased H3K4 methylation and altered gene expression. The KDM5 family normally maintains homeostasis through demethylating H3K4me3/2, thus negatively regulating gene expression. We hypothesised that inhibition of KDM5 may re-establish H3K4 methylation and restore the expression of genes repressed upon loss of KMT2D. Using a selective inhibitor of the KDM5 family we were able to increase H3K4me3 levels in DLBCL cell lines, predominantly at gene promoters, ultimately leading to decreased proliferation and cell death in KMT2D mutant cell lines. This appeared to be driven by an increase in the expression of negative regulators of B cell survival pathways, many of which have previously been described as targets of KMT2D and CREBBP, resulting in diminished BCR signalling. We also observed decreased BCL2 expression in all examined t(14;18) positive cell lines, alongside changes in other BCL2 family members in sensitive cell lines. KDM5 sensitivity was confirmed to be dependent on the presence of KMT2D mutations by generating de novo KMT2D mutant cell lines and correcting naturally mutant cell lines, which displayed greater and lesser sensitivity to KDM5 inhibition respectively. KDM5 inhibition may therefore be an effective therapeutic strategy for ameliorating KMT2D loss of function mutations in malignancies such as germinal centre lymphomas.

overall_design: H3K4me1/H3K4me3 ChIP-seq and RNA-seq were performed in parental WSU-DLCL2 cells, and isogenic KMT2D -/+ mutant generated by CRISPR, treated with DMSO or 1uM KDM5-inh1 for 72h (n=3).

first: J

last: Fitzgibbon

last: Wang


title: The TFIID subunit TAF4 is required for pancreatic beta cell function and identity [single-cell RNA-seq]

city: Strasbourg

first: Irwin

last: Davidson

address: 1 Rue Laurent Fries

country: France

institute: IGBMC

summary: We selectively inactivated the Taf4 subunit of general transcription factor TFIID in adult murine pancreatic beta cells (BCs). Taf4 inactivation rapidly diminishes expression of critical genes involved in BC function leading to increased glycaemia, lowered blood insulin levels, defective glucose-stimulated insulin secretion and in the longer term reduced BC mass through apoptosis of a subpopulation of BCs. Nevertheless, glycaemia and blood insulin levels are stabilised after 11 weeks with mutant animals showing long term survival. Bulk RNA-seq and ATAC-seq and single cell RNA-seq on isolated Langerhans islets show that Taf4 loss leads to major remodelling of chromatin accessibility and gene expression not only in targeted BCs, but also alpha and delta cells. One week after Taf4-loss cells with mixed BC, alpha and/or delta cell identities were observed as well as a population trans-differentiating into alpha cells. Trans-differentiation was associated with the concerted action of the Mafb, Mafg, Bptf and Foxp1 transcription factors. Taf4 is therefore essential for BC function and identity and we identify a novel set of factors associated with BC trans-differentiation.

overall_design: Three single cell RNA-seq samples from isolated Langerhans islets from wild-type control mice or mice where Taf4 has been inactivated in beta cells for 1 or 5 weeks.

first: Thomas

last: Kleiber

first: Guillaume

first: Igor

last: Martianov


title: The TFIID subunit TAF4 is required for pancreatic beta cell function and identity [RNA-seq]

overall_design: 12 total RNA-seq samples from isolated Langerhans islets from wild-type control mice or mice where Taf4 has been inactivated in beta ccells for 1 or 5 weeks.


title: The TFIID subunit TAF4 is required for pancreatic beta cell function and identity


title: An investigation of Burkholderia cepacia complex methylomes via SMRT sequencing and mutant analysis

city: Zürich

first: Gabriella

last: Pessi

address: Winterthurerstrasse 190

institute: University of Zürich

summary: While nucleotide sequences determine an organism’s proteins, methylation of the nucleotides themselves can confer additional properties.  In bacteria, differences in methylation can be used to identify incoming DNA, for example from bacteriophages.  Restriction enzymes detect the methylation status at specific sequence motifs, and cleave improperly methylated, foreign DNA.  Furthermore, methylation of key genetic regions can influence gene expression and hence phenotype.  In this study, we determined the methylated motifs of three strains from the closely-related Burkholderia cepacia complex of opportunistic pathogens.  Three of these motifs were novel, while two are methylated throughout this group of bacteria and beyond and had already been assigned putative causative genes.  To investigate the contribution of the enzymes that methylate and cut DNA, we deleted the genes encoding them from a representative strain.  We found that these genes are important for traits including motility, biofilm formation, cell shape and production of iron acquisition elements.  The mutant strain showed marked defects in replicon partitioning and was more efficient at acquiring foreign DNA, illustrating the importance of methylation in maintaining genome integrity.  Interestingly, we found that one methylase, part of an integrated bacteriophage genome, was essential for maintaining the phage in a dormant state.

overall_design: Burkholderia cepacia complex epigenetics

first: Olga

last: Mannweiler

first: Kirsty

last: Agnoli

first: Marta

last: Pinto-Carbo

first: Martina

last: Lardi

first: Leo

last: Eberl


title: Microfluidic device with brain extracellular matrix promotes structural and functional maturation of human brain organoids

city: Seoul

first: Ann-Na

last: Cho

address: Yonsei-ro 50

country: South Korea

institute: Yonsei university

summary: We established a platform for the brain organoid culture by using human decellularized brain extracellular matrix (BEM) and a microfluidic device. This engineering concept of reconstituting  brain-mimetic microenvironments facilitates development of a reliable culture platform for brain organoids, enabling effective modeling and drug development.

overall_design: Matrigel-encapsulated brain organoid vs BEM-encapsulated brain organoid and conventional plate culture encapsulated with BEM vs microfluidic culture encapsulated with BEM.

first: Seung-Woo


title: The TFIID subunit TAF4 is required for pancreatic beta cell function and identity [ATAC-seq]

overall_design: Three ATAC-seq samples from isolated Langerhans islets from wild-type control mice or mice where Taf4 has been inactivated in beta ccells for 1 or 5 weeks.


title: Ezh2 is essential for the generation of functional yolk sac derived erythro-myeloid progenitors [ATAC-seq]

city: Manchester

first: Georges

last: Lacaud

address: Wilmslow Raod

department: Stem Cell Biology

institute: Cancer Research UK Manchester Institute

summary: Yolk sac (YS) hematopoiesis is critical for the survival of the embryo and a major source of tissue-resident macrophages that persist into adulthood. Here we report that the epigenetic regulator Ezh2 is essential for YS hematopoiesis but dispensable for subsequent aorta–gonad–mesonephros (AGM) blood development. Loss of Ezh2 activity in hemogenic endothelium (HE) leads to the generation of phenotypically intact but functionally deficient erythro-myeloid progenitors (EMP). Ezh2 activity is critical at the onset of the endothelial-to-hematopoietic transition generating EMPs but rapidly dispensable after that. We identified a lack of downregulation of Wnt signaling as the primary reason in the generation of non-functional EMPs. Together, our findings demonstrate a critical and specific role of Ezh2 in modulating Wnt signaling during the generation of EMPs from YS HE.

overall_design: ATAC-seq profile of E10.5 yolk sac erythro-myeloid progenitors

first: Wen

last: Neo

first: Yiran

last: Meng

first: Alba

last: Rodriguez-Meira

first: Emanuele

last: Azzoni

first: Christopher

last: Booth

first: Supat

last: Thongjuea

first: Marella

last: de Bruijn

first: Sten

last: Jacobsen

first: Adam

last: Mead


title: KDM5 inhibition is a novel therapeutic strategy for the treatment of KMT2D mutant lymphomas [1]

first: James

last: Heward

overall_design: The effect of KMD5i was examined on lymphoma cell lines using RNA-seq (n=3, 24 and 72 hours) and H3K4me3 ChIP-seq (n=2, 72 hours). KDM5 isoform expression was also quantified in FL patient samples

first: Jude

last: Fitzgibbon

first: Jun

last: Wang


title: Glomerular endothelial cell mRNA profiles in mice with podocyte depletion

city: New York

first: Weijia

last: Zhang

address: 1 Gustave L. Levy Pl

department: Renal

institute: Icahn School of Medicine at Mount Sinai

summary: we profiled GEC mRNA expression levels in two animal models with podocyte depletion. We used a transgenic mouse model with YFP-labelled GEC for sorting of GEC. Podocyte depletion was induced by injection of mice with a specific anti-podocyte antibody or crossing with a podocyte-specific doxycycline-induced diphtheria toxin A (DTA) expression transgenic mouse model. Both mouse models developed significant proteinuria, glomerulosclerosis, and podocyte depletion.  YFP-positive GECs were sorted from these mice for RNA sequencing. Analysis of the differentially expressed genes (DEGs) between the diseased and control mice revealed significant alteration of metabolism and immune system in the antibody-mediated podocyte depletion model while angiogenesis, vascular development, actin cytoskeleton, and focal adhesion pathways were highly enriched in the DTA model. Apoptosis and cell adhesion pathways were enriched in both models. Apoptosis of GEC was confirmed in the kidney of these mice, which leads to the reduction of GEC number. We also identified a list of genes which were altered in both animal models. Among them, we have further validated Cd63, a molecule which is regulated in human DKD and plays a key role in endothelial cell function. In conclusion, this is the first unbiased approach to determine the genomic-wide response of GEC to podocyte depletion in two different animal models. Our study proves that podocyte depletion could induce GEC injury.

overall_design:
Podocyte depletion was induced by injection of mice with a specific anti-podocyte antibody or crossing with a podocyte-specific doxycycline-induced diphtheria toxin A (DTA) expression transgenic mouse model. YFP-positive GECs were sorted from these mice for RNA sequencing

***Please note that GSE128736 is incorrectly cited for Diabetes 2019, PMID: 31578193***

first: Jia

last: Fu

first: Zhengzi

last: Yi

first: John

last: He


title: Large Drosophila Germline piRNA Clusters are Evolutionarily Labile and Dispensable for Transposon Regulation

status: Public on May 31 2021

first: Daniel

last: Gebert

state: Cambridgeshire

address: Downing Street

department: Department of Genetics

institute: University of Cambridge

type: Non-coding RNA profiling by high throughput sequencing

summary: PIWI proteins and their guiding Piwi-interacting small RNAs (piRNAs) are crucial for fertility and transposon defense in the animal germline. In most species, the majority of piRNAs are produced from distinct large genomic loci, called piRNA clusters. It is assumed that germline-expressed piRNA clusters, particularly in Drosophila, act as master regulators to control the activity of transposons dispersed across the genome. Here, using synteny analysis, we show that large germline clusters are evolutionarily labile, arise at loci characterized by recurrent chromosomal rearrangements, and are mostly species-specific across the Drosophila genus. By engineering chromosomal deletions in D. melanogaster, we demonstrate that the three largest germline clusters, which the generation of >40% of all transposon-targeting piRNAs depend on, are neither required for fertility nor for the regulation of transposon activity in trans. We provide further evidence that dispersed active elements, rather than the regulatory action of large Drosophila germline clusters in trans, may be central for transposon defense.

overall_design: Analysis of effects of germline piRNA cluster deletions on genome, transcriptome and small RNAs

first: Lena

last: Neubert

first: Catrin

last: Lloyd

first: Jinghua

last: Gui

first: Ruth

last: Lehmann

first: Felipe

last: Karam Teixeira


title: Highly Selective Brain-to-Gut Communication via Genetically-Defined Vagus Neurons

city: Charlottesville

middle: N

last: Campbell

state: VA

address: 485 MCCORMICK RD, GILMER HALL ROOM 229 PO BOX 400328

institute: University of Virginia

summary: The vagus nerve innervates many organs, and most if not all of its motor fibers are  cholinergic. However, no one knows its organizing principles; whether or not there are  dedicated neurons with restricted targets that act as "labeled lines" to perform certain  functions – including two opposing ones (gastric contraction versus relaxation). By  performing unbiased transcriptional profiling of DMV cholinergic neurons, we  discovered seven molecularly-distinct subtypes of motor neurons. Then, by using  subtype-specific Cre driver mice, we go on to show that two of these subtypes  exclusively innervate the glandular domain of the stomach where, remarkably, they  contact different enteric neurons releasing functionally opposing neurotransmitters  (acetylcholine versus nitric oxide). Thus, the vagus motor nerve communicates via  genetically-defined labeled lines to control functionally unique enteric neurons within  discrete subregions of the gastrointestinal tract. This discovery provides a conceptual  breakthrough on the division of labor used by the vagus motor nerve to control  function.

overall_design: Single-nuclei RNA-Seq (sNuc-Seq) analysis of 384 samples isolated from the dorsal motor nucleus of the vagus of 12 week old Chat-IRES-cre male mice (C57Bl6/j background)

first: Jenkang

last: Tao

first: Linus

last: Tsai

first: Chen

last: Wu

first: Stephen

last: Liberles

first: Bradford

last: Lowell


title: Sweet corn: low vs. high plant density

city: Urbana

first: Eunsoo

last: Choe

state: Illinois

address: s-306 Turner hall 1102 S. Goodwin Ave.

institute: USDA-ARS

summary: Transcriptional profiling of sweet corn response to plant density (crowding stress). Determine the extent to which hybrid and environment influences crowding stress response and identify crowding stress transcriptional response in sweet corn

overall_design: 2 hybrid x 2 environment x 2 plant density with 4 biological replications

first: Choe

last: Eunsoo

first: Williams

last: Martin II


title: Transcriptomic profiling of oncogenic RIT1 and activated YAP1 in lung epithelial cells

city: Seattle

first: Alice

last: Berger

state: WA

address: 1100 Fairview Ave N

institute: Fred Hutchinson Cancer Center

summary: Based on xenograft data, we have found that YAP1 activation synergizes with RIT1M90I to promote lung cancer. To investigate the molecular basis for the observed functional synergy, we performed whole-genome RNA-seq in SALE cells expressing RIT1M90I and YAP15SA individually or in combination. We find that RIT1M90I and YAP15SA induced very different transcriptional states with little correlation. And, that transcripts altered by RIT1M90I were not enriched for a YAP1 gene signature, whereas YAP15SA drove activation of this signature. In addition, we find that when RIT1M90I and YAP15SA w co-expressed, YAP1-regulated gene expression was enhanced by RIT1M90I compared to YAP15SA alone, with known YAP1 targets such as TNNT2, ITGB2, and COL4A3 showing 2.5-3.6 fold increased expression compared to YAP15SA alone.  These data suggest that RIT1M90I on its own does not trigger YAP1 target transcription but may instead enhance activation of YAP1 targets in settings of YAP1 activation, such as tumors with Hippo pathway loss.

overall_design: Whole transcriptome (mRNA) profiles of 12 samples.  SALE parental, SALE-RIT1M90I, SALE-YAP5SA, and SALE-RIT1M90I + YAP5SA were generated through lentiviral transduction of either, RIT1 M90I,  YAP5SA, or RIT1M90I + YAP5SA). Sequenced in triplicate on Illumina HiSeq2500.

first: Athea

last: Vichas

first: April

last: Lo


title: Transcriptome analysis of genetically engineered murine fusion-negative rhabdomyosarcoma model-derived cell lines

city: Leuven

first: Maurilio

last: Sampaolesi

address: Herestraat 49, ON4 - box 804

department: Department of Development and Regeneration

country: Belgium

institute: KU Leuven

summary: Fusion-negative rhabdomyosarcoma (FN-RMS) is the most common soft tissue sarcoma of childhood arising from undifferentiated skeletal muscle cells. Tumor cells derived from genetically engineered murine models are a valid tool to study cancer biology. In order to investigate the poor myogenic commitment of FN-RMS, we performed RNA sequencing of two cell lines derived from a previously described genetically engineered murine model of FN-RMS. We then performed bioinformatics analysis to compare the transcriptome with murine satellite cells, and identify the key factors that distinguish FN-RMS from skeletal muscle cells.

overall_design:
Bulk RNAseq of murine rhabdomyosarcoma cell lines

This series includes re-analyzed satellite cell samples from GSE103164.

first: Enrico

last: Pozzo

first: N

last: Giarratana

first: G

last: Sassi

first: M

last: Elmastas

first: T

last: Killian

first: C

last: Wang

first: V

last: Marini

first: J

last: Yustein

first: A

last: Uyttebroeck


title: A live single-cell state reporter assay links intra-tumor heterogeneity to metastatic proclivity in Ewing sarcoma

city: Torino

first: Paolo

last: Provero

address: Via Nizza 52

department: Molecular Biotechnology and Health Sciences

country: Italy

institute: University of Turin

summary: Targeting of the most aggressive tumor cell subpopulations is key for effective management of most solid malignancies. However, the metastable nature of tumor heterogeneity, which allows cells to transition between strong and weak tumorigenic phenotypes and the lack of reliable markers of tumor promoting properties hamper identification the most relevant cells. To overcome these obstacles, we designed a functional, cell state-dependent microRNA (miR)-based live cell reporter assay to identify highly tumorigenic cells in xenotransplants of primary Ewing sarcoma (EwS) organoids. Leveraging the inverse relationship between cell pluripotency and miR-145 expression, we successfully separated highly tumorigenic, metastasis-prone (miR-145low) cells from poorly tumorigenic, non-metastatic (miR-145high) counterparts. Gene expression and functional studies of the two cell populations identified EPHB2 receptor as a prognostic biomarker in EwS patients and a major promoter of metastasis. Our study provides a simple and powerful means to identify and isolate tumor cells in states associated with aggressive behavior.

overall_design: Cells from two Ewing sarcoma organoids were sorted into GFP+ and GFP- fractions, corresponding to low- and high-miR-145 expression, respectively. Genes differentially expressed between the two fractions were identified.

first: Tugba

last: Keskin

first: Beatrice

last: Rucci

first: Sandrine

last: Cornaz-Buros

first: Patricia

last: Martin

first: Carlo

last: Fusco

first: Liliane

last: Broye

first: Katarina

last: Cisarova

first: Igor

last: Letovanec

first: Stefano

last: La Rosa

first: Stéphane

last: Cherix

first: Manuel

last: Diezi

first: Raffaele

last: Renella

first: Ivan

last: Stamenkovic

first: Nicolo

last: Riggi


title: Mutant p53 mediates sensitivity to cancer treatment agents in oesophageal adenocarcinoma associated with microRNA and SLC7A11 expression

city: Bedford Park

first: George

last: Mayne

state: SA

address: Room 3D213, Dept of Surgery, Flinders Medical Center

department: Surgery

country: Australia

institute: Flinders University of South Australia

summary: 1) Background: TP53 gene mutations occur in 70% of oesophageal adenocarcinomas (OACs). Given the central role of p53 in controlling cellular response to therapy we investigated the role of mutant (mut-) p53 and SLC7A11 in a CRISPR-mediated JH-EsoAd1 TP53 knockout model. (2) Methods: Response to 2 Gy irradiation, cisplatin, 5-FU, 4-hydroxytamoxifen and endoxifen was assessed, followed by a TaqMan OpenArray qPCR screening for differences in miRNA expression. (3) Results: Knockout of mut-p53 resulted in increased chemo- and radioresistance (2 Gy survival fraction: 38% vs. 56%, p<0.0001) and in altered miRNA expression levels. Target mRNA pathways analyses indicated several potential mechanisms of treatment resistance. SLC7A11 knockdown restored radiosensitivity (2 Gy SF: 46% vs. 73%; p=0.0239), possibly via enhanced sensitivity to oxidative stress. Pathway analysis of the predicted mRNA targets of differentially expressed miRNAs indicated potential involvement in several pathways associated with apoptosis, ribosomes as well as in p53 signalling pathways. (4) Discussion: These results provide a better understanding of the important role of SLC7A11 in cancer metabolism and redox balance and the influence of p53 on these processes. Inhibition of the SLC7A11-gluthatione axis may represent a promising approach to overcome resistance associated with mut-p53.

overall_design: OpenArray high throughput real time PCR of microRNAs --  (i)  triplicate experiments for 3 Test (p53-knockout) and 3 Control (Parental) oesophageal adenocarcinoma cell lines,  (ii)  duplicate experiments for transfection of p53-knockout cells with either an siRNA specific for SLC7A11 or a Control non-binding siRNA.

first: Ann-Kathrin

last: Eichelmann

first: Karen

last: Chiam

first: Steven

middle: L

last: Due

first: Isabell

last: Bastian

first: Frederike

last: Butz

first: Tingting

last: Wang

first: Pamela

last: Sykes

first: Nicholas

last: Clemons

first: David

last: Liu

first: Michael

middle: Z

last: Michael

first: Christos

last: Karapetis

first: Richard

last: Hummel

middle: I

last: Watson

first: Damian

last: Hussey


title: AP-2α Regulates S-phase and is a Marker for Sensitivity to PI3K-inhibitor Buparlisib in Colon Cancer

city: St Louis

first: Tiandao

last: Li

address: 4444 Forest Park Ave

institute: Washington University

overall_design: Refer to individual Series


title: Two distinct mechanisms for X chromosome dosage compensation in Anopheles and Drosophila [RNA-seq]

city: Freiburg

first: Asifa

last: Akhtar

address: Stuebeweg 51

department: Chromatin Regulation

institute: Max Planck Institute of Immunobiology and Epigenetics

summary: Heteromorphic sex chromosomes induce potentially deleterious gene expression imbalances that are frequently corrected by dosage compensation (DC). Three distinct molecular strategies to achieve DC have been previously described in nematodes, fruit flies and mammals. The reason for these mechanistic differences remain unclear: Are they a consequence of distinct genomes and gene content, functional or ecological constraints, or random initial commitment to an evolutionary trajectory? Here, we study DC in the malaria mosquito Anopheles gambiae. The X chromosomes of Anopheles and Drosophila evolved independently, yet from the same ancestral autosome and share a high degree of homology. We find that Anopheles achieves DC by an entirely different mechanism compared to the MSL complex - H4K16ac axis operating in Drosophila. CRISPR knock-out of msl-2 in Anopheles leads to early embryonic lethality and affects both sexes. Transcriptome analyses indicate that this phenotype is not a consequence of defective X chromosome DC, but instead relates to misregulation of developmental genes. Furthermore, Histone H4 Lysine 16 acetylation does not mark an X chromosome territory, neither does it display a sexually dimorphic genome-wide distribution by ChIP. We conclude that a novel pathway confers X chromosome upregulation in male Anopheles. Our findings highlight the pluralism of how organisms cope with gene-dosage alterations and show that different mechanisms can evolve even in scenarios of highly similar genomic and functional constraints.

sample_organism:
Anopheles gambiae
Drosophila melanogaster

overall_design: RNA-seq in msl-2 KO Drosophila melanogaster and Anopheles gambiae.

first: Claudia

last: Valsecchi

first: Gina

last: Renschler


title: Two distinct mechanisms for X chromosome dosage compensation in Anopheles and Drosophila [ChIP-seq]

overall_design: ChIP-seq of H3 and H4K16ac in dissected midguts of male and female Anopheles gambiae.


title: Single-cell RNA-seq of fly glia

city: Houston

first: Hongjie

state: TX

address: one Baylor Plaza

institute: Baylor College of Medicine

summary: Using single-cell RNA-sequencing, we characterized the transcriptomes of Drosophila young and old glial cells from repo+ pan glia and ensheathing glia. scRNAseq was performed using Smart-seq2 platform.

overall_design: Single-cell RNA-seq


title: Transcriptome RNA-Sequencing of regulated genes in Hep3B liver cancer cell by si-SMARCA4

first: Sang Yean

last: Kim

state: Select

address: 222 BANPO-DAERO SEOCHO-GU, SEOUL

institute: The Catholic university of korea

summary: In order to find a new genes regulated by SMARCA4, RNA was extracted 2days after transfection of si-SMARCA4 into a Hep3B cell line. We hypothesized that when SMARCA4 was inhibited, the genes expression will be suppressed.

overall_design: Examination of si-SMARCA4 transfect in Hep3B cell line. RNA-Sequencing was repeated three times..

first: Sang-Yean

first: Suk-Woo

last: Nam


title: Two distinct mechanisms for X chromosome dosage compensation in Anopheles and Drosophila

summary: This SuperSeries is composed of the SubSeries listed below.

platform_organism:
Anopheles gambiae
Drosophila melanogaster


title: In vivo CRISPR screening identifies RNF20/40 as epigenetic regulators of cardiomyocyte maturation

city: Indianapolis

first: Nathan

last: VanDusen

state: IN

department: Pediatrics

institute: Indiana University School of Medicine

summary: Between birth and adulthood cardiomyocytes (CMs) undergo dramatic changes in size, ultrastructure, metabolism, and gene expression, in a process collectively referred to as CM maturation. The transcriptional network that coordinates CM maturation is poorly understood, creating a bottleneck for cardiac regenerative medicine. Forward genetic screens are a powerful, unbiased method to gain novel insights into transcriptional networks, yet this approach has rarely been used in vivo in mammals because of high resource demands. Here we utilized somatic mutagenesis to perform the first reported in vivo CRISPR genetic screen within a mammalian heart. We discovered and validated several novel transcriptional regulators of CM maturation. Among them were RNF20 and RNF40, which form a complex that monoubiquitinates H2B on lysine 120. Mechanistic studies indicated that this epigenetic mark controls dynamic changes in gene expression required for CM maturation. These insights into CM maturation will inform efforts in cardiac regenerative medicine. More broadly, our approach will enable unbiased forward genetics across mammalian organ systems.

overall_design: A CRISPR guide RNA library was recovered from Myh7YFP+ cardiomyocytes, sequenced, and gRNA enrichment calculated. Gene expression was assessed in P6 GATA4/6 knockout cardiomyocytes by RNA-sequencing. Gene expression was assessed in P28 RNF20/40 depleted cardiomyocytes by RNA-sequencing. H2Bub1 ChIP-sequencing was conducted on P1 and P28 cardiomyocytes. H3K4me3 and H3K36me3 ChIP-sequencing was conducted on P7 wildtype and Rnf20 cardiomyocyte-specific KO ventricles.

first: William

last: Pu


title: A549 vs. A549DDP miRNA and mRNA analysis using RNA-seq

city: Monroe

first: George

last: Mattheolabakis

state: LA

address: 1800 Bienville Dr., University of Louisiana at Monroe, room 380

institute: University of Louisiana at Monroe

summary: This study aims to identify pathway involvement in the development of cisplatin (cis-diamminedichloroplatinum (II); CDDP) resistance in A549 lung cancer (LC) cells by utilizing advanced bioinformatics software. We developed CDDP-resistant A549 (A549/DDP) cells through prolonged incubation with the drug and performed RNA-seq on RNA extracts to determined differential mRNA and miRNA expression between A549/DDP and A549 cells

overall_design: Examination of A549 parental cell line vs. A549-cisplatin resistant cell line

first: AKM Nawshad

last: Hossian


title: PRMT5-mediated histone arginine methylation antagonizes transcriptional repression by polycomb complex PRC2 (ChIP-seq)

status: Public on May 30 2021

city: Miami

first: Stephen

last: Nimer

state: FL

address: 1120 NW 14th ST

department: Sylvester Comprehensive Cancer Center

institute: University of Miami

summary: Protein arginine methyltransferase 5 (PRMT5) catalyzes the symmetric di-methylation of arginine residues in histones H3 and H4, marks generally associated with transcriptional repression. However, we found that PRMT5 inhibition (or depletion) led to more genes being down-regulated than up-regulated, indicating that PRMT5 can also act as a transcriptional activator. Indeed, the global level of histone H3K27me3 increases in PRMT5 deficient cells. Although PRMT5 does not directly affect PRC2 enzymatic activity, methylation of histone H3 by PRMT5 abrogates its subsequent methylation by PRC2. Treating AML cells with an EZH2 inhibitor partially restored the expression of approximately 50% of the genes that are initially down-regulated by PRMT5 inhibition, suggesting that the increased H3K27me3 contributes to the transcription repression of these genes. Interestingly, the growth inhibitory effect of PRMT5 inhibition was also partially rescued by combined treatment with EZH2 inhibitor in several leukemia cell lines. Thus, PRMT5-mediated crosstalk between histone marks contributes to its functional effects.

overall_design: Molm13 cells were treated with DMSO (0.05%), EZH2 inhibitor (GSK343) 1 uM or PRMT5 inhibitor (GSK3186000A) 1 uM for 4 days.  DNA was enriched via IP for H3K27me3 and subjected to high throughput sequencing.

first: Fan


title: PRMT5-mediated histone arginine methylation antagonizes transcriptional repression by polycomb complex PRC2 (RNA-seq)

overall_design: Molm13 cells were treated with DMSO, EZH2 inhibitor, PRMT5 inhibitor or combination of both inhibitors for 4 days. Total RNA were purified with RNeasy plus mini kit. Libraries were prepared using standard Illumina protocols with paired RNA sequencing.

last: Liu


title: PRMT5-mediated histone arginine methylation antagonizes transcriptional repression by polycomb complex PRC2


title: Visualizing programmed lineage trajectories at cancer initiation

city: Durham

first: Chaitanya

middle: R

last: Acharya

state: NC

address: 203 Research Drive,Medical Sciences Research Building, Suite 433

department: Dept. of Surgery

institute: Duke University

summary: The sequential accumulation of survival advantage traits – the cancer evolution hypothesis – implies that early detection and treatment of precancers can prevent cancer mortality. In practice, detection and treatment of breast precancer ductal  carcinoma in situ (DCIS) has had minimal impact on reducing breast cancer mortality. The evolution of precancer is not inevitable, suggesting that inciting neoplastic events lead to different trajectories of progression. Here we visualize distinct tumor driver gene programs of lineage specification occurring at cancer initiation. Crainbow fluorescent barcoding of wild-type HER2, d16-HER2, and p95-HER2 enables whole-gland lineage tracing and single-cell reconstruction of the origin of HER2+ breast cancer. Molecular pathology and Crainbow lineage tracing provide genotype:phenotype modules and gene signatures predictive of cancer progression. WT-HER2 neoplasms were rare and proliferative d16-HER2 neoplasms eventually progressed to metastasis, and p95-HER2 induced rapidly invasive and metastatic carcinomas. p95-HER2 carcinomas originate without detectable in situ stages – termed here a minimally detectable “nascent lethal carcinoma”. This nascent lethal program biases the commitment of tumor stem cells toward a metaplastic epithelial-mesenchymal transition-like (EMT) lineage promoting microenvironmental remodeling. Overall, these data suggest that lethality programs begin at initiation, indicating that detection must be improved to identify the nascent lethal lesion.

overall_design: Singe cells obtained from 12 Her2+ tumors that are either d16 or p95.

first: Joshua

last: Ginzel

first: Veronica

last: Lubkov

first: Hidetoshi

last: Mori

first: Peter

middle: G

last: Boone

first: Lauren

last: Rochelle

first: Wendy

last: Roberts

first: Zachary

last: Hartman

first: Erika

last: Crosby

first: Jeffrey

last: Everitt

first: Lawrence

last: Barak

first: Marc

last: Caron

first: Jane

middle: Q

last: Chen

first: Neil

middle: E

last: Hubbard

first: Robert

last: Cardiff

first: Alexander

last: Borowsky

first: H

last: Lyerly

last: Snyder


title: Quality control in scRNA-Seq can discriminate pacemaker cells - the mtDNA bias

status: Public on May 28 2021

city: Dummerstorf

first: Anne-Marie

last: Galow

address: Wilhelm-Stahl-Alle 2

institute: Leibniz-Institute for farm animal biology

summary: Single-cell RNA-Sequencing (scRNA-Seq) provides high-resolution insights into complex tissues. Cardiac tissue, however, poses a major challenge due to the delicate isolation process and the large size of mature cardiomyocytes. Regardless of the experimental technique, captured cells are often impaired and some capture sites may contain multiple or no cells at all. All this refers to “low quality” potentially leading to data misinterpretation. Common standard quality control parameters involve the number of detected genes and transcripts per cell and the fraction of transcripts from mitochondrial genes (%mtDNA). While cutoffs for transcripts and genes per cell are usually user-defined for each experiment or individually calculated, a fixed threshold of 5% mtDNA is standard and often set as default in scRNA-Seq software. However, this parameter is highly dependent on the tissue type. In the heart, mitochondrial transcripts comprise almost 30% of total mRNA due to high energy demands. Here, we demonstrate that a 5%-threshold not only causes an unacceptable exclusion of cardiomyocytes but also introduces a bias that particularly discriminates pacemaker cells. This effect is apparent for our in vitro generated induced-sinoatrial-bodies (iSABs; highly enriched physiologically functional pacemaker cells), and also evident in a public dataset of cells isolated from embryonal murine sinoatrial node tis-sue [2]. Taken together, we recommend omitting this filtering parameter for scRNA-Seq in cardiovascular applications whenever possible.

overall_design: Investigation of subpopulations and their respective features in “induced-sinoatrial-bodies'' (iSABs) assesed by single-cell sequencing using the 10x Genomics system

first: Sophie

last: Kussauer

first: Markus

last: Wolfien

last: David

first: Andreas

last: Hoeflich


title: Murine colon carcinoma cell line (CMT93): control (cont) vs. Lipopolysaccharides (LPS) stimulation

city: Yufu

first: Sotaro

last: Ozaka

state: Oita

address: 1-1 Idaigaoka, Hasama

institute: Oita University

summary: The gene expression profile of mouse colon carcinoma cell line (CMT93) comparing control untreated CMT93 cells with CMT93 cells stimulated with LPS.

overall_design: Two-condition experiment, cont vs. LPS.

first: Akira

last: Sonoda

first: Takashi

last: Kobayashi


title: An unsupervised and broadly applicable method for physical cell interaction profiling of complex tissues

city: Stockholm

first: Martin

last: Enge

address: CCK, Z4

department: Dep of Oncology-Pathology

country: Sweden

institute: Karolinska Institute

summary: Cellular identity in complex multicellular organisms is controlled in part by the physical organization of cells. However, large-scale investigation of the cellular interactome has been technically challenging. Here we develop Cell Interaction by Multiplet sequencing (CIM-seq), an unsupervised and high-throughput method to analyze direct physical cell-cell interactions between every cell types presented in a tissue. CIM-seq is based on RNA sequencing of incompletely dissociated cells, followed by computational deconvolution into constituent cell types using machine learning. Contrary to previous deconvolution-based methods, CIM-seq estimates parameters such as number of cells and cell types in each multiplet directly from sequencing the data, making it compatible with high-throughput droplet-based methods. When applied to gut epithelium, or whole dissociated lung and spleen, CIM-seq correctly identifies known interactions, including those between different cell lineages and immune cells. In the colon, CIM-seq identifies a previously unrecognized goblet cell subtype expressing the wound-healing marker Plet1, which is directly adjacent to colonic stem cells. Our results from different tissue types demonstrate that CIM-seq is broadly applicable to profile cell type interactions in different tissue types using in an unsupervised manner.

overall_design: Deconvolution of transcriptional profiles from cell clusters of 2 or more cells (multiplets) identifying their constituent cell types.

first: Nathanael

last: Andrews

first: Natalie

last: Geyer

first: Ninib

last: Baryawno

first: Marco

last: Gerling


title: RNA-seq analysis of MCF7 cells with/without  silencing TRPS1/HDAC2

city: Nanjing

first: liming

last: chen

state: Jiangsu

address: Qixiaqu wenyuanlu 1hao

department: college of life science

institute: Nanjing Normal University

summary: We reported the transcriptional regulation function and mechanism of TRPS1.

overall_design: Examination of transcription repression role of TRPS1 in breast cancer cells by RNA-seq in duplicates using illumina hiseq2500.

first: Liming

last: Chen


title: Distinct nuclear compartment-associated genome architecture in the developing mammalian brain

city: San Francisco

first: Tomasz

last: Nowakowski

state: Ca

institute: UCSF

summary: Nuclear compartments are thought to play a role in three-dimensional genome organization and gene expression. In mammalian brain, the architecture and dynamics of nuclear compartment-associated genome organization is not known. In this study, we developed Genome Organization using CUT and RUN Technology (GO-CaRT) to map genomic interactions with two nuclear compartments—the nuclear lamina and nuclear speckles—from different regions of the developing mouse, macaque and human brain. Lamina-associated domain (LAD) architecture in cells in vivo is distinct from that of cultured cells, including major differences in LADs previously considered to be cell type invariant. In the mouse and human forebrain, dorsal and ventral neural precursor cells have differences in LAD architecture that correspond to their regional identity. LADs in the human and mouse cortex contain transcriptionally highly active sub-domains characterized by broad depletion of histone-3-lysine-9 dimethylation. Evolutionarily conserved LADs in human, macaque and mouse brain are enriched for transcriptionally active neural genes associated with synapse function. By integrating GO-CaRT maps with genome-wide association study data, we found speckle-associated domains to be enriched for schizophrenia risk loci, indicating a physical relationship between these disease-associated genetic variants and a specific nuclear structure. Our work provides a framework for understanding the relationship between distinct nuclear compartments and genome function in brain development and disease.

sample_organism:
Macaca mulatta
Homo sapiens
Mus musculus

overall_design: RNA-seq data of E13 mouse cortex and subpallium; LaminB GO-CaRT data of E13 mouse cortex and subpallium NPCs, E13 liver, E13 lung, V-SVZ cultured NSCs derived from postnatal day 7, E80 macaque cortex and subpallium/MGE, NIH/3T3 cells, HEK293T cells; SON GO-CaRT of HEK293T cells; H3K9me2 CUT&RUN of E13 mouse cortex and NIH/3T3 cells; H3K9me3 CUT&RUN of NIH/3T3 cells; H3K27me3 CUT&RUN of NIH/3T3 cells.

first: Ryan

last: Delgado


title: Muscle Transcriptional Networks Linked to Resistance Exercise Training Hypertrophic Response Heterogeneity

city: Birmingham

first: Kaleen

middle: M

last: Lavin

state: Alabama

address: 908 20th Street South, Room 201-C

department: Medicine

institute: University of Alabama at Birmingham

summary: Bulk polyA-RNAseq was performed on vastus lateralis muscle samples from older adults ≥65y at baseline and post-14 wk resistance exercise training (high-low-high intensity). The primary study used network biology algorithms to create gene networks of highly connected transcripts, and then linked these to change in muscle size induced by exercise training.

overall_design:
Vastus lateralis samples from males and females ≥65y were collected at baseline [time 1] (n=31) and post-RT [time 3] (n=22).

This series includes re-analyzed samples from GSE157585.

first: Margaret

middle: B

last: Bell

first: Jeremy

last: McAdam

first: Bailey

last: Peck

first: R

last: Walton

first: Samuel

last: Windham

first: S

last: Tuggle

first: Douglas

last: Long

first: Philip

last: Kern

first: Charlotte

last: Peterson

first: Marcas

last: Bamman


title: A Single Neuron Scale Molecular Phenotype Map of the Right Atrial Ganglionic Plexus in the Pig Heart

city: Philadelphia

first: Rajanikanth

last: Vadigepalli

state: PA

address: 1020 Locust St, 314C

department: Pathology

institute: Jefferson University

summary: We developed a spatially-tracked single neuron transcriptomics map of an intrinsic cardiac ganglion - the right atrial ganglionic plexus (RAGP) that is a critical mediator of vagal control of the sinoatrial node (SAN) activity. We developed a 3D representation of RAGP with extensive mapping of neurons and used neuronal tracing to identify the spatial distribution of the subset of neurons that project to the SAN. RNAseq of laser capture microdissected neurons revealed a distinct composition of RAGP neurons compared to CNS neuronal subtypes. High-throughput qPCR of hundreds of laser capture microdissected single neurons led to a surprising finding that cholinergic and catecholaminergic neuronal markers Th and Chat were correlated, suggesting multipotential phenotypes that can drive neuroplasticity within RAGP. Interestingly, no single gene or module was an exclusive marker of RAGP neuronal connectivity to SAN. Neuropeptide-receptor coexpression analysis revealed a combinatorial paracrine neuromodulatory network within RAGP, informing follow-on studies on the vagal control of RAGP to regulate cardiac function in health and disease.

overall_design: 142 regional neuronal samples from the Right Atrial Ganglionic Plexus (RAGP) of a female Yucatan minipig were collected through Laser Capture Microdissection (LCM) and subjected to single-cell scale RNAseq using the Smart-3SEQ protocol. After quality check we were left with 90 samples with detectable expression in approximately 15,000 genes.

first: Alison

last: Moss

first: Shaina

last: Robbins

first: Sirisha

last: Achanta

first: Lakshmi

last: Kuttippurathu

first: Scott

last: Turick

first: Sean

last: Nieves

first: Peter

last: Hanna

first: Jeffrey

last: Ardell

first: Kalyanam

last: Shivkumar

first: James

middle: S

last: Schwaber


title: Single-nucleus RNA sequencing of deep and superficial sublayers of hippocampal area CA1 in epilepsy

city: San Joan d'Alacant

first: Jose

middle: P

last: Lopez-Atalaya

address: Av Ramon y Cajal s/n

department: Instituto Neurociencias

country: Spain

institute: CSIC-UMH

summary: We performed gene expression profiling of FACS purified nuclei from hippocampal area CA1 from adult male mice (Calb1xTdtomato mice) subjected to kainic acid-induced status epilepticus (chronic epileptic phase) and control animals.

overall_design: Young adult male mice (6-7 weeks old) were subjected to kainic acid-induced status epilepticus or sham control (n = 2/group). Sequencing of single nuclei was performed after 12 weeks post-status when animals were in the chronic epileptic phase.

first: Angel

last: Marquez-Galera

first: Liset

last: Menendez de la Prida


title: Single cell gene expression analysis of neurons in the right atrial ganglionic plexus in the pig

summary: As part of the NIH SPARC program efforts to study the intrinsic cardiac nervous system, we developed the first comprehensive molecular phenotyping data on the right atrial ganglionated plexus (RAGP) within the intrinsic cardiac nervous system of the pig. We collected hundreds of single neurons from the RAGP and assayed these for expression of a wide range of genes relevant to neuronal functions. The sinoatrial node (SAN) of 4 pigs were injected with FastBlue, a retrograde tracer, labeling neurons projecting to the SAN.  The RAGP of those pigs were then collected and sectioned.  Both SAN-projecting and non-SAN-projecting single neurons in RAGP were collected through laser capture microdissection (LCM), allowing for spatial mapping of all collected samples. Over 400 single neurons were collected (n=4 animals) and each neuron was assayed for over 300 genes using high throughput microfluidic qPCR using BioMark. Neuronal phenotypes were distinguished by multivariate analysis showing patterns of network activity between multiple genes. Interestingly, these phenotypes show no spatial preferences within the RAGP.  Our results revealed extensive combinatorial expression of neurotransmitters across the RAGP neuronal phenotypes. Additionally, there was a large overlap in expression profiles of SAN-projecting and non-SAN-projecting neurons, without any single gene or module acting as a distinguishing marker between the connectionally labeled groups. Previous studies have focused on cholinergic and catecholaminergic processes, showing evidence for extensive protein expression of cholinergic markers in RAGP.  Our findings are in stark contrast to these results and demonstrate high gene expression correlation between cholinergic and catecholaminergic markers such as tyrosine hydroxylase (Th) and choline acetyltransferase (ChAT) in single neurons. This finding suggests that cells are poised for a rapid shift in neurotransmitter signaling as well as the possibility for post transcriptional regulation of neuron phenotype plasticity. Our present findings significantly expand the list of neuromodulatory peptides and their receptors known to be expressed in RAGP. For example, neuropeptide Y and somatostatin showed distinct co-expression patterns with their respective receptors with significant non-overlapping expression of transmitters and cognate receptors across single neurons. Taken together, our results reveal a complex organization of neuronal networks within RAGP-SAN complex that is not entirely governed by connectivity or spatial location, with combinatorial patterns of local paracrine networks with potential influence on cardiac function.

overall_design:
Over 400 single neurons were collected (n=4 animals) and each neuron was assayed for over 200 genes using high throughput microfluidic qPCR using BioMark.
Twenty one 96 × 96 BioMark Arrays were used to measure gene expression across the (422 samples before QC) single-cell samples. The same serial dilution sample set was included in each chip to verify reproducibility and test for technical variability. Samples from each animal were run across three chips to obtain data on 283 genes per sample. Each set of chip runs for a given animal contained overlapping assays that served as technical replicates to evaluate chip-to-chip variability.
Data Normalization: Individual qRT-PCR results were examined to determine the quality of the qRT-PCR based on melt-curve analysis. Following this initial quality control, samples with >30% failed reactions and genes with >20% failed reactions were excluded from present analysis. A further 10 samples were determined to be outliers due overall gene expression distributions and were removed from the present analysis. Upon filtering based on these criteria, a total of 405 single-cell samples (152 non SAN-projecting neurons, 169 SAN-projecting neurons, and 84 neurons without reliable connectivity information) and 241 different gene assays were carried forward in the present analysis, with 211 genes showing >60% detectable expression across all RAGPs. Raw Ct values for individual samples were normalized against a median expression level of a subset of 140 robustly expressed genes (genes with greater than 60% working reactions) across all animals to obtain -DCt values.The vector of median sample expression value was chosen over potential reference genes based on comparison of stable expression across all samples against known housekeeping genes using the ‘selectHKs’ function in the NormqPCR package in R. The following equation was used to calculate -DCt values for each gene: -DCtgene = (median sample expression) - Ctgene.
The -DCt data were then rescaled using the median across all samples within a gene using the following equation:
-DDCtgene=-(DCtsample- DCtacross-sample-median).


title: Single-cell Analysis of Surface Topography-induced Foreign Body Response to Silicone Implants

first: H. Courtney

last: Hodges

address: 1 Baylor Plaza, BCM 229

department: Department of Molecular & Cellular Biology

summary: Silicone-based medical devices are widely used in chronic implants and are generally perceived to be safe. However, immune-related complications including malignancies have recently been linked to textured breast implants. Here, we examine the influence of clinically approved breast implants surface features on host immune responses. Prosthetics with surface roughness of 0, 4, and 90 (Ra) were implanted in mammary fat pads of mice for 2 weeks and cells adjacent to the resulting tissue capsules were evaluated for foreign body immune responses using single-cell RNA-seq. Our findings identify a unique and finely tuned surface topography that is capable of modulating implant immunity to suppress foreign body response.

overall_design: Single-cell 5' RNA-seq (scRNA-seq) of immune cells derived from fibrotic capsules of C57BL/6 mice with silicone implants. All samples are pooled from 5 independent mice in the same condition and processed using 10x 5' scRNA-seq according to the manufacturer's instructions.

first: Joshua

middle: C

last: Doloff

first: Omid

last: Veiseh

first: H

first: Robert

last: Langer


title: ATAC-Seq of foetal blood progenitors

city: Edinburgh

first: Katrin

last: Ottersbach

address: 5 Little France Drive

department: Centre for Regenerative Medicine

institute: University of Edinburgh

summary: Chromatin accessibility was assessed by ATAC-Seq in lymphoid-primed multipotent progenitors (LMPPs) from human foetal livers (FLs) and mouse wild-type FLs as well as FLs from mouse embryos that express the oncofusion Mll-AF4 in the definitive blood system. The aim of this study was to establish whether overall chromatin accessibility at key haematopoietic sites and loci that have been linked to leukaemia are differentially accessible in human vs mouse LMPPs and whether this is altered by the expression of the Mll-AF4 oncofusion.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: 1000-5000 LMPPs (Lineage- CD34+ CD38- CD19- CD45RA+) from human FLs (10-20 weeks of gestation) and 5000-15000 LMPPs (Lin- CD45+ ckit+ Sca1+ Flt3+) from wild-type and Mll-AF4-expressing E14.5 FLs were isolated. Cells were sorted straight into transposition buffer, followed by tagmentation, PCR amplification and library clean up

first: Eric

last: Antunes

first: Camille

last: Malouf

first: Michael

last: O'Dwyer

first: Abdenour

last: Soufi


title: Adult mouse LN, dermis and epidermis Treg scRNAseq

status: Public on May 29 2021

first: Margaret

last: Lowe

state: CA

address: 513 Parnassus Avenue, HSW1201B

summary: Comparison of regulatory T cell transcriptional signatures in adult mouse lymph node, dermis, and epidermis

overall_design: 1 sample of adult murine back epidermal Treg, 1 sample of adult murine dermal Treg, 1 sample of adult murine lymph node Tregs

last: Moreau

first: Jarish

last: Cohen

middle: D

last: Rosenblum